Mammalian Inscuteable Regulates Spindle Orientation and Cell Fate in the Developing Retina

Žigman, Mihaela; Cayouette, Michel; Charalambous, Christoforos; Schleiffer, Alexander; Hoeller, Oliver; Dunican, Dara J.; McCudden, Christopher R.; Firnberg, Nicole; Barres, Ben A.; Siderovski, David P.; Knoblich, Juergen A.

doi:10.1016/j.neuron.2005.09.030

Cited by 122 publications

(118 citation statements)

References 33 publications

(39 reference statements)

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“…Once all binding sites on LGN are occupied by mInsc, spindle orientation should be random. In support of this hypothesis, horizontal mitotic spindles are enriched during the early stages of neurogenesis when mInsc expression is low (28). More importantly, the enrichment of horizontally oriented mitotic spindles persists during the later neurogenic stages of neurogenesis in mInsc knock-out mice, whereas it is less pronounced in wild-type mice.…”

Section: Discussionmentioning

confidence: 66%

Analysis and modeling of mitotic spindle orientations in three dimensions

Jüschke

Xie

Postiglione

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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The orientation of the mitotic spindle determines the relative size and position of the daughter cells and influences the asymmetric inheritance of localized cell fate determinants. The onset of mammalian neurogenesis, for example, coincides with changes in spindle orientation. To address the functional implications of this and related phenomena, precise methods for determining the orientation of the mitotic spindle in complex tissues are needed. Here, we present methodology for the analysis of spindle orientation in 3D. Our method allows statistical analysis and modeling of spindle orientation and involves two parameters for horizontal and vertical bias that can unambiguously describe the distribution of spindle orientations in an experimental sample. We find that 3D analysis leads to systematically different results from 2D analysis and, surprisingly, truly random spindle orientations do not result in equal numbers of horizontal and vertical orientations. We show that our method can describe the distribution of spindle orientation angles under different biological conditions. As an example of biological application we demonstrate that the adapter protein Inscuteable (mInsc) can actively promote vertical spindle orientation in apical progenitors during mouse neurogenesis.

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Section: Discussionmentioning

confidence: 66%

Analysis and modeling of mitotic spindle orientations in three dimensions

Jüschke

Xie

Postiglione

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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“…Insc was first identified in larval Drosophila neuroblasts as a partner of Par3 (Bazooka in flies) colocalizing with polarity proteins right after delamination (7)(8)(9), and later shown to bind Pins (10). Mammalian Insc homologues endowed with similar properties have been discovered in rat retina (11) and in mouse developing skin (12). Insc overexpression is sufficient to induce apico-basal divisions in cells that normally divide in a planar fashion such as Drosophila embryonic ephitelial cells (7), vertebrate neuroepithelial progenitors (13), and mouse skin progenitors (6).…”

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confidence: 99%

Inscuteable and NuMA proteins bind competitively to Leu-Gly-Asn repeat-enriched protein (LGN) during asymmetric cell divisions

Culurgioni

Alfieri

Pendolino

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Coupling of spindle orientation to cellular polarity is a prerequisite for epithelial asymmetric cell divisions. The current view posits that the adaptor Inscuteable (Insc) bridges between Par3 and the spindle tethering machinery assembled on NuMA∶LGN∶Gαi GDP , thus triggering apico-basal spindle orientation. The crystal structure of the Drosophila ortholog of LGN (known as Pins) in complex with Insc reveals a modular interface contributed by evolutionary conserved residues. The structure also identifies a positively charged patch of LGN binding to an invariant EPE-motif present on both Insc and NuMA. In vitro competition assays indicate that Insc competes with NuMA for LGN binding, displaying a higher affinity, and that it is capable of opening the LGN conformational switch. The finding that Insc and NuMA are mutually exclusive interactors of LGN challenges the established model of force generators assembly, which we revise on the basis of the newly discovered biochemical properties of the intervening components.A symmetric cell divisions regulate the position and the fate choice of daughter cells, with impact on numerous phenotypes of multicellular organisms. During development, asymmetric divisions coordinate cell growth with cell specification to determine tissue morphogenesis, while in adult life they sustain tissue homeostasis and regeneration (1). In asymmetric divisions specific cortical landmarks instruct the orientation of the mitotic spindle to promote unequal partitioning of fate determinants in cellular systems as diverse as Caenorhabditis elegans zygotes, Drosophila neuroblasts, as well as vertebrate skin and neural progenitors (2). Spindle coupling to polarity cues involves the recruitment at cortical sites of molecular devices, known as force generators, whose main task is to capture astral microtubules emanating from the spindle poles and to establish pulling forces. Core components of force generators are the evolutionary conserved NuMA∶LGN∶Gαi complexes, termed Mud∶Pins∶Gαi in flies. Topologically, tetratricopeptide repeats (TPR) present in the N-terminal portion of LGN mediate the interactions with NuMA, while GoLoco motifs at the C terminus serve as a docking platform for four Gαi GDP subunits anchored at the plasma membrane via a myristoyl group (3).LGN exclusively binds to GDPloaded Gαi (4). The association of cortical NuMA with the microtubule motor Dynein/Dynactin (5) provides a sliding anchorage for depolymerizing microtubules, whose shrinkage pulls towards the cortex the connected spindle pole. FRET studies revealed that LGN behaves as a conformational switch held in a closed form in interphase by head-to-tail interactions (3).An issue intimately related to how force generators are assembled is how they are recruited at sites of polarization. In polarized asymmetric divisions, the apico-basal polarity axis is established by the asymmetrical distribution of Par3∶Par6∶aPKC at the apical cortex, which are able to recruit NuMA∶LGN∶Gαi GDP via an adaptor named Inscuteable (Insc) (6). Insc w...

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“…Ainsi, en favorisant l'augmentation localisée des sous-unités G liées au GDP, GPR-1 et GPR-2, celles-ci favorisent l'augmentation des forces exercées sur les microtubules. À quelques différences près, les mécanismes moléculaires en cause dans le positionnement du fuseau mitotique chez C. elegans sont conservés dans les neuroblastes de drosophile (Figure 2B) [19,20]), ainsi que dans les progéniteurs neuronaux de mammifères se divisant de manière asymétrique [21,22]. Cependant, le rôle de ces machineries moléculaires dans la division asymétrique d'autres types cellulaires, en l'occurrence ceux de mammifères, ainsi que leurs effecteurs en aval, restent encore à caractériser.…”

Section: Positionnement Asymétrique Du Fuseau Mitotiqueunclassified